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1.
Anticancer Drugs ; 33(8): 741-751, 2022 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-35946523

RESUMO

The oncogenic role of ST8SIA6-AS1 in different cancers was reported, including hepatocellular carcinoma (HCC). However, the underlying mechanism has not been completely explored. Real time quantitative PCR analysis was conducted to assess the ST8SIA6-AS1, miR-651-5p and TM4SF4 expression in HCC tissues and cells. Cell counting kit-8 and wound-healing migration assays were adopted to evaluate the HCC cell proliferation and migration, respectively. The expression of apoptosis-related proteins (Bax and Bcl-2) in human colorectal cancer cells (HCC) was determined by western blotting. In addition to bioinformatics analysis, RNA immunoprecipitation studies and luciferase reporter assays were undertaken to investigate the direct target relationship among ST8SIA6-AS1 and miR-651-5p or TM4SF4. Highly expressed ST8SIA6-AS1 and TM4SF4 as well as poorly expressed miR-651-5p were detected in HCC tissues and cells. Clinically, miR-651-5p expression in HCC tissues is negatively correlated with ST8SIA6-AS1 or TM4SF4. Cell functional assays demonstrated that ST8SIA6-AS1 silencing resulted in weakened proliferative and migratory capacities in HCC cells in addition to increase Bax expression and reduced Bcl-2 expression. ST8SIA6-AS1 exhibited its oncogenic function by sponging tumor suppressor miR-651-5p, and the anti-oncogenic of miR-651-5p was offset by its TM4SF4. The manipulation of ST8SIA6-AS1/miR-651-5p/TM4SF4 axis-mediated oncogenicity in HCC might shed new light on HCC diagnosis and therapy.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , RNA Longo não Codificante , Carcinogênese/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Glicoproteínas de Membrana/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Sialiltransferases/genética , Sialiltransferases/metabolismo , Proteína X Associada a bcl-2/metabolismo
2.
Glycobiology ; 32(9): 736-742, 2022 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-35789385

RESUMO

The ST6GAL1 Golgi sialyltransferase is upregulated in many human malignancies, however, detection of ST6GAL1 protein in cancer tissues has been hindered by the prior lack of antibodies. Recently, numerous commercial antibodies for ST6GAL1 have become available, however, many of these do not, in fact, recognize ST6GAL1. Decades ago, the CD75 cell-surface epitope was mistakenly suggested to be the same molecule as ST6GAL1. While this was rapidly disproven, the use of CD75 as a synonym for ST6GAL1 has persisted, particularly by companies selling "ST6GAL1" antibodies. CD75 is reportedly a sialylated epitope which appears to encompass a range of glycan structures and glycan carriers. In this study, we evaluated the LN1 and ZB55 monoclonal antibodies, which are advertised as ST6GAL1 antibodies but were initially developed as CD75-recognizing antibodies (neither was raised against ST6GAL1 as the immunogen). Importantly, the LN1 and ZB55 antibodies have been widely used by investigators, as well as the Human Protein Atlas database, to characterize ST6GAL1 expression. Herein, we used cell and mouse models with controlled expression of ST6GAL1 to compare LN1 and ZB55 with an extensively validated polyclonal antibody to ST6GAL1. We find that LN1 and ZB55 do not recognize ST6GAL1, and furthermore, these 2 antibodies recognize different targets. Additionally, we utilized the well-validated ST6GAL1 antibody to determine that ST6GAL1 is overexpressed in bladder cancer, a finding that contradicts prior studies which employed LN1 to suggest ST6GAL1 is downregulated in bladder cancer. Collectively, our studies underscore the need for careful validation of antibodies purported to recognize ST6GAL1.


Assuntos
Neoplasias da Bexiga Urinária , Animais , Antígenos CD/metabolismo , Epitopos , Humanos , Camundongos , Polissacarídeos , Sialiltransferases/metabolismo
3.
ACS Synth Biol ; 11(8): 2837-2845, 2022 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-35802806

RESUMO

3'-Sialyllactose (3'-SL) is one of the most important and simplest sialylated human milk oligosaccharides. In this study, a plasmid-based pathway optimization along with chromosomal integration strategies was applied for 3'-SL production. Specifically, the precursor CMP-Neu5Ac synthesis pathway genes and α2,3-sialyltransferase-encoding gene were introduced into Escherichia coli BL21(DE3)ΔlacZ to realize 3'-SL synthesis. Genes nanA and nanK involved in Neu5Ac catabolism were further deleted to reduce the metabolic flux of competitive pathway. Several α2,3-sialyltransferases from different species were selected to evaluate the sialylation effect. The precursor pools were balanced and improved by optimizing key enzyme expression involved in the UDP-GlcNAc and CMP-Neu5Ac synthesis pathway. Finally, an additional α2,3-sialyltransferase expression cassette was integrated into chromosome to maximize 3'-SL synthesis, and 4.5 g/L extracellular 3'-SL was produced at a shake-flask level. The extracellular 3'-SL concentration was raised to 23.1 g/L in a 5 L bioreactor fermentation, which represents the highest extracellular value ever reported.


Assuntos
Escherichia coli , Sialiltransferases , Escherichia coli/metabolismo , Humanos , Leite Humano/metabolismo , Oligossacarídeos/metabolismo , Sialiltransferases/genética , Sialiltransferases/metabolismo
4.
Environ Toxicol ; 37(10): 2398-2411, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35730485

RESUMO

BACKGROUND: Triple-negative breast cancer (TNBC), the most aggressive subtype of breast cancer, always exhibits a poor prognosis due to high risk of early recurrence and distant metastasis. Long noncoding RNAs (lncRNAs) have been reported as crucial regulators in breast cancer. However, the functions and action mechanisms of lncRNA ST8SIA6-AS1 in TNBC are largely unknown. METHODS: Quantitative real-time PCR and western blot assays were used to measure the expression levels of different genes and proteins. Cell proliferation ability was monitored by CCK-8, colony forming and flow cytometry assays. Wound healing and transwell assays were performed to evaluate cell migration and invasion. The regulatory mechanisms of ST8SIA6-AS1 in TNBC were confirmed by dual luciferase reporter and RIP assays. A mouse xenograft model was established to investigate the role of ST8SIA6-AS1 in TNBC tumor growth. RESULTS: ST8SIA6-AS1 displayed a higher expression in TNBC cells. Silencing ST8SIA6-AS1 impaired cell proliferation, cell cycle progression, migration, and invasion in vitro, and slowed tumor growth in vivo. Mechanistically, ST8SIA6-AS1 could facilitate the expression of its target CDCA3 (cell division cycle associated protein 3) and inactivate the p53/p21 signaling by inhibiting miR-145-5p. Moreover, miR-145-5p exerted a tumor-suppressive activity by targeting CDCA3. The tumor-suppressive effects induced by ST8SIA6-AS1 knockdown were abated by the down-regulation of miR-145-5p or the up-regulation of CDCA3. CONCLUSION: ST8SIA6-AS1 exerts an oncogenic role in TNBC by interacting with miR-145-5p to up-regulate CDCA3 expression and inactivate the p53/p21 signaling, highlighting ST8SIA6-AS1 as a promising molecular target to combat TNBC.


Assuntos
MicroRNAs , RNA Longo não Codificante , Neoplasias de Mama Triplo Negativas , Animais , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Sialiltransferases/genética , Sialiltransferases/metabolismo , Transdução de Sinais/genética , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
5.
Glycobiology ; 32(8): 701-711, 2022 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-35661210

RESUMO

Interaction of immune cells with the systemic environment is necessary for the coordinated development and execution of immune responses. Monocyte-macrophage lineage cells reside at the junction of innate and adaptive immunity. Previously we reported that the sialyltransferase ST6GAL1 in the extracellular milieu modulates B cell development and IgG production, granulocyte production, and attenuates acute airway inflammation to bacterial challenge in mouse models. Here, we report that extracellular ST6GAL1 also elicits profound responses in monocyte-macrophage lineage cells. We show that recombinant ST6GAL1 adheres to subsets of thioglycolate-elicited inflammatory cells in the mouse peritoneum and to cultured human monocyte THP-1 cells. Exposure of the inflammatory cells to recombinant ST6GAL1 elicited wholesale changes in the gene expression profile of primary mouse myeloid cells; most notable was the striking up-regulation of monocyte-macrophage and monocyte-derived dendritic cell development pathway signature genes and transcription factors PU.1, NFκB and their target genes, driving increased monocyte-macrophage population and survival ex vivo. In the cultured human monocyte cells, the essential cell surface receptor of the monocyte-macrophage lineage, the M-CSF receptor (M-CSF-R, Csfr1) was a target of extracellular ST6GAL1 catalytic activity. Extracellular ST6GAL1 activated the M-CSF-R and initiated intracellular signaling events, namely, the nuclear translocation of NFκB subunit p65, and phosphorylation of ERK 1/2 and AKT. The findings implicate extracellular ST6GAL1 in monocyte development by a mechanism initiated at the cell surface and support an emerging paradigm of an extracellular glycan-modifying enzyme as a central regulator coordinating immune hematopoietic cell development and function.


Assuntos
Fator Estimulador de Colônias de Macrófagos , Monócitos , Animais , Antígenos CD/metabolismo , Diferenciação Celular , Humanos , Fator Estimulador de Colônias de Macrófagos/metabolismo , Macrófagos/metabolismo , Camundongos , Monócitos/metabolismo , Fosforilação , Sialiltransferases/genética , Sialiltransferases/metabolismo , Transdução de Sinais , Células THP-1
6.
Glycobiology ; 32(9): 803-813, 2022 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-35746897

RESUMO

The glycosylation of immunoglobulin G (IgG) has attracted increased attention due to the impact of N-glycan modifications at N297 on IgG function, acting primarily through modulation of Fc domain conformation and Fcγ receptor-binding affinities and signaling. However, the mechanisms regulating IgG glycosylation and especially α2,6-sialylation of its N-glycan remain poorly understood. We observed previously that IgG is normally sialylated in mice with B cells lacking the sialyltransferase ST6Gal1. This supported the hypothesis that IgG may be sialylated outside of B cells, perhaps through the action of hepatocyte-released plasma ST6Gal1. Here, we demonstrate that this model is incorrect. Animals lacking hepatocyte expressed ST6Gal1 retain normal IgG α2,6-sialylation despite the lack of detectable ST6Gal1 in plasma. Moreover, we confirmed that B cells were not a redundant source of IgG sialylation. Thus, while α2,6-sialylation is lacking in IgG from mice with germline ablation of ST6Gal1, IgG α2,6-sialylation is normal in mice lacking ST6Gal1 in either hepatocytes or B cells. These results indicate that IgG α2,6-sialylation arises after release from a B cell but is not dependent on plasma-localized ST6Gal1 activity.


Assuntos
Imunoglobulina G , Sialiltransferases , Animais , Glicosilação , Imunoglobulina G/genética , Imunoglobulina G/metabolismo , Camundongos , Polissacarídeos/química , Receptores de IgG , Sialiltransferases/genética , Sialiltransferases/metabolismo
7.
Oxid Med Cell Longev ; 2022: 9381203, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35498127

RESUMO

Long noncoding RNA taurine-upregulated gene 1 (TUG1) is considered to be involved in postischemic cerebral inflammation, whereas polysialic acid (polySia, PSA), the product of St8sia2, constitutes polysialylated neural adhesion cell molecule (PSA-NCAM) in both mice and humans and that cerebral PSA-NCAM level is elevated in neuronal progenitor cells in response to transient focal ischemia. Herein, we aim to identify novel miRNAs that bridge the functions of St8sia2 and TUG1 in ischemia-associated injuries. In both in vivo (C57BL/6J mouse ischemia/reperfusion, I/R model) and in vitro (mouse neuroblastoma N2A cell oxygen glucose deprivation/reoxygenation, OGD model) settings, we observed upregulated TUG1 and St8sia2 after the induction of ischemic injury, accompanied by reduced miR-3072-3p expression. We performed siRNA-induced TUG1 knockdown combined with the induction of ischemic injury; the results showed that inhibiting TUG1 expression led to the reduced infarct area and improved neurological deficit. Through bioinformatics analysis, miR-3072-3p was found to target both St8sia2 and TUG1, which was subsequently verified by the luciferase reporter system and RNA binding protein immunoprecipitation assay. Also, the addition of miR-3072-3p mimic/inhibitor resulted in reduced/elevated St8sia2 expression at the protein level. Further studies revealed that in both in vivo and in vitro settings, TUG1 bound competitively to miR-3072-3p to regulate St8sia2 expression and promote apoptosis. In summary, targeting the TUG1/miR-3072-3p/St8sia2 regulatory cascade, a novel cascade we identified in cerebral ischemia injury, may render feasible therapeutic possibilities for overcoming cerebral ischemic insults.


Assuntos
MicroRNAs , RNA Longo não Codificante/genética , Traumatismo por Reperfusão , Animais , Infarto Cerebral , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Traumatismo por Reperfusão/genética , Sialiltransferases , Taurina
8.
Int J Mol Sci ; 23(9)2022 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-35563598

RESUMO

Schizophrenia is a serious psychiatric disorder that affects the social life of patients. Psychiatric disorders are caused by a complex combination of genetic (G) and environmental (E) factors. Polysialylation represents a unique posttranslational modification of a protein, and such changes in neural cell adhesion molecules (NCAMs) have been reported in postmortem brains from patients with psychiatric disorders. To understand the G × E effect on polysialylated NCAM expression, in this study, we performed precise measurements of polySia and NCAM using a disrupted-in-schizophrenia 1 (DISC1)-mutant mouse (G), a mouse model of schizophrenia, under acute stress conditions (E). This is the first study to reveal a lower number and smaller length of polySia in the suprachiasmatic nucleus of DISC1 mutants relative to those in wild-type (WT) mice. In addition, an analysis of polySia and NCAM responses to acute stress in five brain regions (olfactory bulb, prefrontal cortex, suprachiasmatic nucleus, amygdala, and hippocampus) revealed that the pattern of changes in these responses in WT mice and DISC1 mutants differed by region. These differences could indicate the vulnerability of DISC1 mutants to stress.


Assuntos
Proteínas do Tecido Nervoso , Esquizofrenia , Sialiltransferases , Animais , Encéfalo/metabolismo , Humanos , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Moléculas de Adesão de Célula Nervosa/genética , Moléculas de Adesão de Célula Nervosa/metabolismo , Córtex Pré-Frontal/metabolismo , Esquizofrenia/genética , Esquizofrenia/metabolismo , Sialiltransferases/metabolismo
9.
Int J Mol Sci ; 23(10)2022 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-35628171

RESUMO

Gangliosides (glycosphingolipids containing one or more sialic acids) are highly expressed in neural tissues in vertebrates, and four species (GM1a, GD1a, GD1b, GT1b) are predominant in mammalian brains. GM3 is the precursor of each of these four species and is the major ganglioside in many nonneural tissues. GM3 synthase (GM3S), encoded by ST3GAL5 gene in humans, is a sialyltransferase responsible for synthesis of GM3 from its precursor, lactosylceramide. ST3GAL5 mutations cause an autosomal recessive form of severe infantile-onset neurological disease characterized by progressive microcephaly, intellectual disability, dyskinetic movements, blindness, deafness, intractable seizures, and pigment changes. Some of these clinical features are consistently present in patients with ST3GAL5 mutations, whereas others have variable expression. GM3S knockout (KO) mice have deafness and enhanced insulin sensitivity, but otherwise do not display the above-described neurological defects reported in ST3GAL5 patients. The authors present an overview of physiological functions and pathological aspects of gangliosides based on findings from studies of GM3S KO mice and discuss differential phenotypes of GM3S KO mice versus human GM3S-deficiency patients.


Assuntos
Surdez , Epilepsia , Sialiltransferases , Animais , Surdez/enzimologia , Modelos Animais de Doenças , Epilepsia/enzimologia , Humanos , Camundongos , Camundongos Knockout , Sialiltransferases/deficiência , Sialiltransferases/metabolismo
10.
Blood Adv ; 6(13): 3945-3955, 2022 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-35507766

RESUMO

Although the sialyltransferases ST3GAL1 and ST3GAL2 are known to transfer sialic acid to the galactose residue of type III disaccharides (Galß1,3GalNAc) in vitro, sialylation of O-linked glycosylated proteins in living cells has been largely attributed to ST3GAL1. To examine the role of ST3GAL2 in O-sialylation, we examined its expression during differentiation of human-induced pluripotent stem cells (iPSCs) into hematopoietic progenitor cells (HPCs) and megakaryocytes (MKs). ST3GAL1 and ST3GAL2 each became highly expressed during the differentiation of iPSCs to HPCs but decreased markedly in their expression upon differentiation into MKs, suggesting coordination of expression during megakaryopoiesis. To further delineate their role in these processes, we generated ST3GAL1-, ST3GAL2-, and doubly deficient human iPSC lines. Binding of the peanut agglutinin lectin, which reports the presence of unsialylated Galß1,3GalNAc glycan chains, was strongly increased in HPCs and MKs derived from double-knockout iPSCs and remained moderately increased in cells lacking either one of these sialyltransferases, demonstrating that both can serve as functional cellular O-glycan sialyltransferases. Interestingly, the HPC markers CD34 and CD43, as well as MK membrane glycoprotein (GP) GPIbα, were identified as major GP substrates for ST3GAL1 and ST3GAL2. In contrast, O-sialylation of GPIIb relied predominantly on the expression of ST3GAL2. Finally, although disruption of ST3GAL1 and ST3GAL2 had little impact on MK production, their absence resulted in dramatically impaired MK proplatelet formation. Taken together, these data establish heretofore unknown physiological roles for ST3GAL1 and ST3GAL2 in O-linked glycan sialylation in hemato- and megakaryocytopoiesis.


Assuntos
Megacariócitos , Sialiltransferases/metabolismo , Diferenciação Celular , Humanos , Polissacarídeos , Especificidade por Substrato
11.
Biochem Biophys Res Commun ; 608: 52-58, 2022 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-35390672

RESUMO

Sialylation, the final stage of post-translational modification of proteins, is achieved in the Golgi apparatus and is related to the malignant phenotype of cancer. Disialylation of ganglioside (GD3) by St8sia1 and polysialylation by St8sia2 and 4 have been shown to be related to malignant phenotypes; however, di/oligosialylation by St8sia6 is still unknown. In this study, we analyzed the malignant phenotype of St8sia6 and found that upregulation of St8sia6 in melanoma B16 cells increased anchorage-independent cell growth, which was not due to sialic acid cleavage by a sialidase. Moreover, unlike other sialyltransferases, St8sia6 localized to the endoplasmic reticulum (ER). We found that the localization to the Golgi apparatus could be regulated by swapping experiments using St8sia2; however, the malignant phenotype did not change. These data demonstrate that the enhancement of anchorage-independent cell growth by St8sia6 is not due to its localization of ER, but is due to the expression of the protein itself.


Assuntos
Retículo Endoplasmático , Neoplasias , Sialiltransferases , Processos de Crescimento Celular , Retículo Endoplasmático/enzimologia , Retículo Endoplasmático/metabolismo , Gangliosídeos/metabolismo , Complexo de Golgi/metabolismo , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Sialiltransferases/metabolismo
12.
Bioconjug Chem ; 33(5): 773-780, 2022 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-35426312

RESUMO

Tools to interrogate glycoconjugate-protein interactions in the context of living cells are highly attractive for the identification of critically important functional binding partners of glycan-binding proteins. These interactions are challenging to study due to the low affinity and rapid dissociation rates of glycan-protein binding events. The use of photo-cross-linkers to capture glycan-protein interaction complexes has shown great promise for identifying binding partners involved in these interactions. Current methodologies use metabolic oligosaccharide engineering (MOE) to incorporate photo-cross-linking sugars. However, these MOE strategies are not amenable to all cell types and can result in low incorporation and cell-surface display of the photo-cross-linking probe, limiting their utility for studying many types of interactions. We describe here an exo-enzymatic strategy for selectively introducing photo-cross-linking probes into cell-surface glycoconjugates using the recombinant human sialyltransferase ST6GAL1 and a diazirine-linked CMP-Neu5Ac derivative. Probe introduction is highly efficient, amenable to different cell types, and resulted in improved cross-linking when compared to MOE. This exo-enzymatic labeling approach can selectively introduce the photo-cross-linking sugar onto specific glycan epitopes and subclasses by harnessing the specificity of the sialyltransferase employed, underscoring its potential as a tool to interrogate and identify glycoconjugate ligands for diverse glycan-binding proteins.


Assuntos
Diazometano , Sialiltransferases , Reagentes de Ligações Cruzadas/química , Diazometano/química , Glicoconjugados/química , Humanos , Polissacarídeos/química , Proteínas/química
13.
Int J Cancer ; 151(6): 957-966, 2022 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-35467766

RESUMO

Cancer patients treated with capecitabine and oxaliplatin (XELOX) often develop hand-foot syndrome (HFS) or palmar-plantar erythrodysesthesia. Genetic variation in ST6GAL1 is a risk factor for type-2 diabetes (T2D), a disease also associated with HFS. We analysed genome-wide association data for 10 toxicities in advanced colorectal cancer (CRC) patients from the COIN and COIN-B trials. One thousand and fifty-five patients were treated with XELOX ± cetuximab and 745 with folinic acid, fluorouracil and oxaliplatin ± cetuximab. We also analysed rs6783836 in ST6GAL1 with HFS in CRC patients from QUASAR2. Using UK Biobank data, we sought to confirm an association between ST6GAL1 and T2D (17 384 cases, 317 887 controls) and analysed rs6783836 against markers of diabetes, inflammation and psoriasis. We found that 68% of patients from COIN and COIN-B with grade 2-3 HFS responded to treatment as compared to 58% with grade 0-1 HFS (odds ratio [OR] = 1.1, 95% confidence interval [CI] = 1.02-1.2, P = 2.0 × 10-4 ). HFS was also associated with improved overall survival (hazard ratio = 0.92, 95% CI = 0.84-0.99, P = 4.6 × 10-2 ). rs6783836 at ST6GAL1 was associated with HFS in patients treated with XELOX (OR = 3.1, 95% CI = 2.1-4.6, P = 4.3 × 10-8 ) and was borderline significant in patients receiving capecitabine from QUASAR2, but with an opposite allele effect (OR = 0.66, 95% CI = 0.42-1.03, P = .05). ST6GAL1 was associated with T2D (lead SNP rs3887925, OR = 0.94, 95% CI = 0.92-0.96, P = 1.2 × 10-8 ) and the rs6783836-T allele was associated with lowered HbA1c levels (P = 5.9 × 10-3 ) and lymphocyte count (P = 2.7 × 10-3 ), and psoriasis (P = 7.5 × 10-3 ) beyond thresholds for multiple testing. In conclusion, HFS is a biomarker of treatment outcome and rs6783836 in ST6GAL1 is a potential biomarker for HFS with links to T2D and inflammation.


Assuntos
Antígenos CD , Capecitabina , Síndrome Mão-Pé , Oxaliplatina , Sialiltransferases , Antígenos CD/genética , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Capecitabina/efeitos adversos , Cetuximab/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/genética , Fluoruracila , Variação Genética , Estudo de Associação Genômica Ampla , Síndrome Mão-Pé/genética , Humanos , Inflamação/complicações , Oxaliplatina/efeitos adversos , Psoríase/genética , Sialiltransferases/genética
14.
Bioconjug Chem ; 33(5): 781-787, 2022 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-35437982

RESUMO

Glycan binding often mediates extracellular macromolecular recognition events. Accurate characterization of these binding interactions can be difficult because of dissociation and scrambling that occur during purification and analysis steps. Use of photocrosslinking methods has been pursued to covalently capture glycan-dependent interactions in situ; however, use of metabolic glycan engineering methods to incorporate photocrosslinking sugar analogs is limited to certain cell types. Here, we report an exo-enzymatic labeling method to add a diazirine-modified sialic acid (SiaDAz) to cell surface glycoconjugates. The method involves the chemoenzymatic synthesis of diazirine-modified CMP-sialic acid (CMP-SiaDAz), followed by sialyltransferase-catalyzed addition of SiaDAz to desialylated cell surfaces. Cell surface SiaDAzylation is compatible with multiple cell types and is facilitated by endogenous extracellular sialyltransferase activity present in Daudi B cells. This method for extracellular addition of α2-6-linked SiaDAz enables UV-induced crosslinking of CD22, demonstrating the utility for covalent capture of glycan-mediated binding interactions.


Assuntos
Diazometano , Ácido N-Acetilneuramínico , Diazometano/química , Glicoproteínas/química , Ácido N-Acetilneuramínico/química , Polissacarídeos/química , Ácidos Siálicos/química , Sialiltransferases/química
15.
J Immunol ; 208(8): 1845-1850, 2022 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-35379746

RESUMO

Inhibitory receptors have a critical role in the regulation of immunity. Siglecs are a family of primarily inhibitory receptors expressed by immune cells that recognize specific sialic acid modifications on cell surface glycans. Many tumors have increased sialic acid incorporation. Overexpression of the sialyltransferase ST8Sia6 on tumors led to altered immune responses and increased tumor growth. In this study, we examined the role of ST8Sia6 on immune cells in regulating antitumor immunity. ST8Sia6 knockout mice had an enhanced immune response to tumors. The loss of ST8Sia6 promoted an enhanced intratumoral activation of macrophages and dendritic cells, including upregulation of CD40. Intratumoral regulatory T cells exhibited a more inflammatory phenotype in ST8Sia6 knockout mice. Using adoptive transfer studies, the change in regulatory T cell phenotype was not cell intrinsic and depended on the loss of ST8Sia6 expression in APCs. Thus, ST8Sia6 generates ligands for Siglecs that dampen antitumor immunity.


Assuntos
Neoplasias , Sialiltransferases , Animais , Camundongos , Camundongos Knockout , MicroRNAs/genética , MicroRNAs/imunologia , Ácido N-Acetilneuramínico/imunologia , Neoplasias/imunologia , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/imunologia , Sialiltransferases/genética , Sialiltransferases/imunologia
16.
J Dent Res ; 101(8): 942-950, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-35238237

RESUMO

Mucin in saliva plays a critical role in the hydration and lubrication of the oral mucosa by retaining water molecules, and its impaired function may be associated with hyposalivation-independent xerostomia. Age-dependent effects on salivary gland function and rheological properties of secreted saliva are not fully understood as aging is a complex and multifactorial process. We aimed to evaluate age-related changes in the rheological properties of saliva and elucidate the underlying mechanism. We performed ex vivo submandibular gland (SMG) and sublingual gland (SLG) perfusion experiments to collect saliva from isolated glands of young (12 wk old) and aged (27 mo old) female C57BL/6J mice and investigate the rheological properties by determining the spinnbarkeit (viscoelasticity). While fluid secretion was comparable in SMG and SLG of both mice, spinnbarkeit showed a significant decrease in SLG saliva of aged mice than that of young mice. There were no significant differences in GalNAc concentration between young and aged SLG saliva. Liquid chromatography/tandem mass spectrometry analysis of SLG saliva revealed that (Hex)1 (HexNAc)1 (NeuAc)1 at m/z 793.31 was the most abundant O-glycan structure in SLG saliva commonly detected in both mice. Lectin staining of salivary gland tissue showed that SLG stained strongly with Maackia amurensis lectin II (MAL II) while Sambucus nigra agglutinin (SNA) stained little, if any, SLG. The messenger RNA expression of St3gal1 that encodes an α-2,3 sialic acid sialyltransferase SIAT4-A showed a decrease in SLG of aged mice, confirmed by a Western blot analysis. Lectin blot analysis in SLG saliva revealed that the relative signal intensity detected by MAL II was significantly lower in aged SLG. Our results suggest that spinnbarkeit decreases in SLG of aging mice due to downregulation of sialic acid linked to α-2,3 sialic acid sialyltransferase expression.


Assuntos
Glândula Sublingual , Xerostomia , Envelhecimento , Animais , Feminino , Lectinas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Ácido N-Acetilneuramínico/metabolismo , Saliva/metabolismo , Sialiltransferases , Glândula Submandibular/metabolismo , Xerostomia/metabolismo
17.
Cell ; 185(7): 1172-1188.e28, 2022 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-35303419

RESUMO

Intestinal mucus forms the first line of defense against bacterial invasion while providing nutrition to support microbial symbiosis. How the host controls mucus barrier integrity and commensalism is unclear. We show that terminal sialylation of glycans on intestinal mucus by ST6GALNAC1 (ST6), the dominant sialyltransferase specifically expressed in goblet cells and induced by microbial pathogen-associated molecular patterns, is essential for mucus integrity and protecting against excessive bacterial proteolytic degradation. Glycoproteomic profiling and biochemical analysis of ST6 mutations identified in patients show that decreased sialylation causes defective mucus proteins and congenital inflammatory bowel disease (IBD). Mice harboring a patient ST6 mutation have compromised mucus barriers, dysbiosis, and susceptibility to intestinal inflammation. Based on our understanding of the ST6 regulatory network, we show that treatment with sialylated mucin or a Foxo3 inhibitor can ameliorate IBD.


Assuntos
Microbioma Gastrointestinal , Doenças Inflamatórias Intestinais , Sialiltransferases/genética , Animais , Homeostase , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Camundongos , Muco/metabolismo , Sialiltransferases/metabolismo , Simbiose
18.
J Biol Chem ; 298(4): 101726, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35157848

RESUMO

Heterogeneity within the glycocalyx influences cell adhesion mechanics and signaling. However, the role of specific glycosylation subtypes in influencing cell mechanics via alterations of receptor function remains unexplored. It has been shown that the addition of sialic acid to terminal glycans impacts growth, development, and cancer progression. In addition, the sialyltransferase ST6Gal-I promotes epidermal growth factor receptor (EGFR) activity, and we have shown EGFR is an 'allosteric mechano-organizer' of integrin tension. Here, we investigated the impact of ST6Gal-I on cell mechanics. Using DNA-based tension gauge tether probes of variable thresholds, we found that high ST6Gal-I activity promotes increased integrin forces and spreading in Cos-7 and OVCAR3, OVCAR5, and OV4 cancer cells. Further, employing inhibitors and function-blocking antibodies against ß1, ß3, and ß5 integrins and ST6Gal-I targets EGFR, tumor necrosis factor receptor, and Fas cell surface death receptor, we validated that the observed phenotypes are EGFR-specific. We found that while tension, contractility, and adhesion are extracellular-signal-regulated kinase pathway-dependent, spreading, proliferation, and invasion are phosphoinositide 3-kinase-Akt serine/threonine kinase dependent. Using total internal reflection fluorescence microscopy and flow cytometry, we also show that high ST6Gal-I activity leads to sustained EGFR membrane retention, making it a key regulator of cell mechanics. Our findings suggest a novel sialylation-dependent mechanism orchestrating cellular mechanics and enhancing cell motility via EGFR signaling.


Assuntos
Neoplasias Ovarianas , Sialiltransferases , Linhagem Celular Tumoral , Movimento Celular , Receptores ErbB/metabolismo , Feminino , Humanos , Integrinas/metabolismo , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/fisiopatologia , Fosfatidilinositol 3-Quinases/metabolismo , Sialiltransferases/metabolismo
19.
Biomed Khim ; 68(1): 7-17, 2022 Jan.
Artigo em Russo | MEDLINE | ID: mdl-35221292

RESUMO

Sialic acids (SA) are derivatives of neuraminic acid; they are located at the terminal position in the chains of monosaccharide residues of various glycoconjugates. SA play a dual role, they either mask recognition sites, or, on the contrary, represent biological targets that can be recognized by receptor proteins and serve as ligands. The desialylation/sialylation processes can be viewed as a dynamic modification regulated by sialyltransferases and sialidases in response to external or internal stimuli. This review describes the structural and functional diversity and the potential use of SA fractions as biomarkers for various pathological conditions. Almost any extreme effects on the body and inflammatory processes lead to an increase in the level of both total and free SA in the blood and tissues. Possible reasons for the increase of sialoglycoconjugate metabolism indicators in biological material include activation of the hepatocyte synthesis and secretion of various acute-phase proteins, many of which are sialoglycoproteins, violation of the membrane integrity and destruction of body cells, and also high activity of sialidases (neurominidases) and sialyltransferases. Most acute and chronic liver diseases are characterized by the decrease in the total level of SA in the blood serum (because many plasma proteins are synthesized and glycosylated in hepatocytes). Aberrant sialylation results in changes of sialoglycoconjugate structure, its ability to perform biological functions and half-life. Glycosylation is the most common post-translational modification of proteins in the virus, which not only promotes the formation of specific conformation of viral proteins, but also modulates their interaction with receptors and affects host cell recognition, viral replication and infectivity. Serum total SA concentration increases in some benign and inflammatory conditions, which indicates a lack of specificity and limits their use for early detection and screening of neoplastic diseases. Nevertheless, determining blood SA level and measuring concentration of existing biomarkers can be used to improve diagnostic indicators, to stage and monitor therapeutic response in some types of cancer, when the need for specificity is less than for diagnosis. Clinical and diagnostic value of determining the sialoglycoconjugate metabolic indicators, including changes in the content of both SA fractions and specific proteins in various biological fluids and tissues, lies in establishing the causes and mechanisms of biochemical changes in the body in certain diseases.


Assuntos
Neoplasias , Ácidos Siálicos , Glicoconjugados , Humanos , Neoplasias/metabolismo , Neuraminidase/metabolismo , Ácidos Siálicos/metabolismo , Sialiltransferases/metabolismo
20.
Transl Psychiatry ; 12(1): 51, 2022 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-35115485

RESUMO

Altered long-range connectivity is a common finding across neurodevelopmental psychiatric disorders, but causes and consequences are not well understood. Genetic variation in ST8SIA2 has been associated with schizophrenia, autism, and bipolar disorder, and St8sia2-/- mice show a number of related neurodevelopmental and behavioral phenotypes. In the present study, we use conditional knockout (cKO) to dissect neurodevelopmental defects and behavioral consequences of St8sia2 deficiency in cortical interneurons, their cortical environment, or in the di- and mesencephalon. Neither separate nor combined cortical and diencephalic ablation of St8sia2 caused the disturbed thalamus-cortex connectivity observed in St8sia2-/- mice. However, cortical ablation reproduced hypoplasia of corpus callosum and fornix and mice with di- and mesencephalic ablation displayed smaller mammillary bodies with a prominent loss of parvalbumin-positive projection neurons and size reductions of the mammillothalamic tract. In addition, the mammillotegmental tract and the mammillary peduncle, forming the reciprocal connections between mammillary bodies and Gudden's tegmental nuclei, as well as the size of Gudden's ventral tegmental nucleus were affected. Only mice with these mammillary deficits displayed enhanced MK-801-induced locomotor activity, exacerbated impairment of prepulse inhibition in response to apomorphine, and hypoanxiety in the elevated plus maze. We therefore propose that compromised mammillary body connectivity, independent from hippocampal input, leads to these psychotic-like responses of St8sia2-deficient mice.


Assuntos
Corpos Mamilares , Sialiltransferases , Animais , Corpos Mamilares/fisiologia , Mesencéfalo , Camundongos , Tegmento Mesencefálico
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